During embryogenesis, cells become specified to assume a particular developmental fate according to their position in the embryo and as a result of interactions with their neighboring cells. These processes are controlled by genetic programs that involve differential expression of regulatory genes in a precisely controlled temporal and spatial pattern. One group of embryonic cells, the mesoderm, invaginates into the interior of the embryo during gastrulation, and subsequently develops into a complex pattern of different muscle types, the heart, and other tissues. The overall objective of the proposed research is to study the mechanisms that lead to the specification of individual cell identities in this germ layer. In the fruit fly Drosophila, these mechanisms can be analyzed with a powerful combination of genetic, molecular and biochemical methods. Three previously identified homeo box genes, tinman (tin), bagpipe (bap) and S59 will be used as paradigms to study the regulation of mesoderm and muscle development. Our genetic analysis suggests that tin and bap are the key genes that specify dorsal mesodermal cells to develop into gut musculature and the heart. Thus, the fundamental subdivision of the mesoderm into a dorsal (visceral) and ventral (somatic) primordium is ultimately a result of the restriction of tin and bap expression to the dorsal cells of the mesoderm. One focus of our research will therefore be to analyze the mechanisms involved in the spatial regulation of these genes. Genetic experiments will be used to examine whether inductive signals from the epidermis influence the spatial expression of tin and bap. The cis-regulatory sequences required for tin and bap regulation will be determined in transgenic flies, and biochemical experiments will be performed to characterize trans-acting factors. To obtain insight into the functions of tin and bap, the interaction with potential downstream genes will be examined. The S59 expression pattern suggests a function of this gene to specify the identity of particular body wall muscles, and this will be tested through the identification and analysis of S59 mutants. Because of the growing evidence that many regulatory mechanisms in development are conserved during evolution, the results of this research are expected to allow a better understanding of mesoderm, muscle and heart formation also in vertebrates and humans.